title: HIV-1 genotyping of the protease-reverse transcriptase and integrase genes to detect mutations that confer antiretroviral resistance.
abstract: Major advances in antiretroviral (ARV) therapy during the last decade have made HIV-1 infections a chronic, manageable disease. In spite of these significant advancements, ARV drug resistance remains a hurdle for HIV-infected patients who are committed to lifelong treatments. Several commercially marketed and/or laboratory-developed tests (LDT) are available to detect resistance-associated mutations (RAMs) in HIV-1, by genotyping. These genotyping tests mainly comprise polymerase chain reaction (PCR)-amplification and population, nucleotide sequencing (Sanger methodology) of a large part of the protease (PR), reverse transcriptase (RT), and integrase (IN) genes. In this chapter, we describe HIV-1 PR, RT, and IN genotyping on clinical samples (plasma), using the LDT methodology performed at Janssen Diagnostics BVBA, Belgium (JDx), where the PR-RT genotyping is used as input, to generate a CE-marked vircoTYPE™ HIV-1 report while the IN genotyping is performed as a research-use-only (RUO) assay. The complete HIV-1 PR gene (297 bp; 99 amino acids) and a large part of the RT gene (the first 1,200 bp; 400 amino acids) are amplified and sequenced as a single 1,497 bp fragment. Genotyping of the IN gene is performed by amplification and sequencing of the RT-IN region (the last 459 bp; 153 amino acids of RT with the complete 867 bp; 289 amino acids of IN). This methodology allows identification of nucleoside/-nucleotide reverse transcriptase, non-nucleoside reverse transcriptase, protease, and integrase inhibitor (NRTI, NtRTI, NNRTI, PI, INI) RAMs in the PR-RT and IN genes, which allows to predict viral response against current ARV regimens.
authors.affiliation: Janssen Diagnostics BVBA, Beerse, Belgium.
authors.forename: Peter
authors.initials: P
authors.lastname: Van den Eede
authors.forename: Liesbeth
authors.initials: L
authors.lastname: Van Wesenbeeck
authors.forename: Yvan
authors.initials: Y
authors.lastname: Verlinden
authors.forename: Maxim
authors.initials: M
authors.lastname: Feyaerts
authors.forename: Veerle
authors.initials: V
authors.lastname: Smits
authors.forename: Ann
authors.initials: A
authors.lastname: Verheyen
authors.forename: Leen
authors.initials: L
authors.lastname: Vanhooren
authors.forename: Alain
authors.initials: A
authors.lastname: Deloof
authors.forename: Jorge
authors.initials: J
authors.lastname: Villacian
authors.forename: Theresa
authors.initials: T
authors.lastname: Pattery
mesh_terms: D019295:Computational Biology / Q000379:methods; D024882:Drug Resistance, Viral / Q000235:genetics*; D005838:Genotype; D060005:Genotyping Techniques / Q000379:methods; D019427:HIV Integrase / Q000235:genetics* / Q000378:metabolism; D016333:HIV Protease / Q000235:genetics* / Q000378:metabolism; D054303:HIV Reverse Transcriptase / Q000235:genetics* / Q000378:metabolism; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics*; D006801:Humans; D012367:RNA, Viral / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA
chemical_list: D012367:RNA, Viral; D019427:HIV Integrase; D054303:HIV Reverse Transcriptase; D016333:HIV Protease